The present invention relates to peptides capable of regulating various physiological functions of Fibroblast Growth Factor-5 (hereinafter, referred to as xe2x80x9cFGF-5xe2x80x9d), and to a pharmaceutical composition containing the peptide as an active component.
FGF-5 is a secretory protein whose expression can be observed in a central nervous system. FGF-5 is known to have physiological functions such as fibroblast proliferation promoting activity and fibroblast mutating activity. FGF-5 is also known to change the proliferation ability of an NIH3T3 fibroblast cell transformed by introducing FGF-5 gene under the control of a permanent expression promoter. The transformed cell secretes FGF-5 into a culture supernatant, which strikingly promotes proliferation of Balb/c3T3 (one of other kinds of fibroblast) and a vascular endothelial cell. FGF-5 polypeptide expressed by introducing an FGF-5 gene expression plasmid into E.coli also remarkably promotes the proliferation of Balb/c3T3 and a vascular endothelial cell.
FGF-5 is also known for its activity as a neurotrophic factor for supporting the survival of neurocyte. FGF-5 is also expressed in skeletal muscle cells. Both of FGF-5 contained in a skeletal muscle cell extract and FGF-5 expressed by introducing FGF-5 gene expression plasmid in E.coli are known to remarkably promote the survival of cultured motoneurons. This fact strongly suggests that FGF-5 is a neurotrophic factor for motoneurons. FGF-5 is also known to be expressed in mouse and rat brains. From an experiment of a primary culture of a cranial nerve cell, FGF-5 is considered to behave as a neurotrophic factor for cholinergic and serotonergic neurocytes in brain.
Analysis of recently-developed FGF-5 knockout mouse suggests that FGF-5 is involved in a hair growth cycle. Most components effective for promoting or inhibiting the development or growth of a head or body hair are conventionally obtained as synthetic compounds or by screening natural substances derived from plants or microorganisms for those having the activity by using model animals. However, since nutritional environment of the hair root has been the prior consideration in developing hair development promoting or inhibiting agent or a hair growth agent which contains the above-mentioned active component, most studies focused on anti-inflammation or disinfection of the skin, inhibition of androgen, and improvement or inhibition of the blood circulation of the treated point or the environmental conditions of the hair root. Thus, no attention has been paid so far to the principle of the mechanism of an animal hair generation, and accordingly there may be a chance that FGF-5 involved in the control of the hair growth cycle may be used to obtain a hair development or hair growth regulating agent based on the mechanism of an animal hair generation.
FGF-5 protein is known to have a structure of serially linked Exons 1, 2 and 3, which is obtained by transcribing an FGF-5-protein-coding gene to mRNA and splicing the transcript to form mature mRNA comprising serially linked Exons 1, 2 and 3. The translation frame starts from the translation starting codon ATG (which encodes methionine) in Exon 1, across Exon 2 to the stop codon in Exon 3, and produces a protein having 268 amino acids (in the case of human) or a protein having 264 amino acids (in the case of mouse) [Zhan X. et al., Mol. Cell. Biol., Vol. 8, pp. 3487-3495 (1988); Haub, O. et al., Proc. Natl. Acad., Sci., USA, Vol. 87, pp. 8022-8026 (1990)]. Furthermore, FGF-5S, an alternatively spliced form of FGF-5 lacking Exon 2 was found, which was selectively expressed in brain and skin of human and mouse and the structure thereof have been determined [Ozawa, K. et al., J. Biol. Chem., Vol.273 (44), pp.29262-29271 (1998) and Japanese Patent Application Laid-Open No. 8-75994]. FGF-5S by itself exerted a weak activity as a neurotrophic factor while it also exhibited an activity as an antagonist of FGF-5 [Ozawa, K. et al, J. Biol. Chem., Vol. 273 (44), pp. 29261-29271 (1998)].
Since FGF-5 has various functions such as promoting cellular proliferation, supporting survival and differentiation of neurocytes and controlling hair growth cycle, there has been a demand for finding and studying a substance for controlling the physiological functions of FGF-5.
Platelets play an important role in the hemostatis of organisms, and formation of thrombus. Accordingly, there has been a demand for a substance for increasing the number of platelets, which may be used as a drug for regulating the platelets to counter thrombocytopenia resulting from the side-effects of chemotherapy of cancer or from serious infectious disease. Platelets and parents of platelets (i.e., megakaryocytes) are formed through differentiation, proliferation and maturation of a pluripotent blood stem cell via the action of a hematopoietic factor. Thrombopoietin has recently been reported as the hematopoietic factor involved in the differentiation to give megakaryocytes. However, since administration of thrombopoietin has also been reported to induce antibody production, thrombopoietin is presently considered not suitable as a platelet increasing substance. Accordingly, there has also been a demand for finding and studying a low-molecular controlling substance that has a differentiating activity to give the megakaryocyte without any side-effect.
The present inventors have gone through intensive study to solve the above-described problems, regarding amino acid sequences of FGF-5 and FGF-5S (i.e., the alternatively spliced form of FGF-5), and found that a peptide fragment (hereinafter, referred to as a xe2x80x9cFGF-5 peptidexe2x80x9d) partially containing one of these amino acid sequences had an ability of controlling physiological functions of FGF-5, thereby accomplishing the present invention.
Thus, the present invention provides either: (a) a peptide comprising the entire amino acid sequence shown in SEQ ID NO: 1, or a continuous part of the amino acid sequence shown in SEQ ID NO: 1; or (b) a peptide comprising the amino acid sequence shown in SEQ ID NO: 1 optionally added with 1 to 10 amino acids at the both ends, and having an activity of controlling a physiological function of FGF-5.
Furthermore, the present invention provides either: (a) a peptide comprising the entire amino acid sequence shown in SEQ ID NO: 2, or a continuous part of the amino acid sequence shown in SEQ ID NO: 2; or (b) a peptide comprising the amino acid sequence shown in SEQ ID NO: 2 optionally added with 1 to 10 amino acids at the both ends, and having an activity of controlling a physiological function of FGF-5.
The present invention also provides a peptide of any one of: (a) a peptide comprising the amino acid sequence shown in SEQ ID NO: 3; (b) a peptide comprising an amino acid sequence having one amino acid substitution in the amino acid sequence shown in SEQ ID NO: 3, and having an activity of controlling a physiological function of FGF-5; or (c) a peptide comprising the amino acid sequence shown in SEQ ID NO: 3 optionally added with 1 to 10 amino acids at the N-terminus thereof, and having an activity of controlling a physiological function of FGF-5.
The present invention also provides a peptide of any one of: (a) a peptide comprising the amino acid sequence shown in SEQ ID NO: 4; (b) a peptide comprising an amino acid sequence having one amino acid substitution in the amino acid sequence shown in SEQ ID NO: 4, and having an activity of controlling a -physiological function of FGF-5; or (c) a peptide comprising the amino acid sequence shown in SEQ ID NO: 4 optionally added with 1 to 10 amino acids at the N-terminus thereof, and having an activity of controlling a physiological function of FGF-5.
Examples of the physiological function of FGF-5 include regulating hair development or hair growth, regulating nutrition or function of cranial nerve system, regulating platelets, and promoting or inhibiting proliferation or differentiation of a vascular endothelial cell, a fibroblast, a myoblast, a chondrocyte, an osteoblast or a glia cell.
The present invention further provides a pharmaceutical composition comprising any one of the above-described peptides.
This specification includes all or part of the contents as disclosed in the specifications of Japanese Patent Application Nos. 11-88364 and 2000-6643, which are priority documents of the present application.